Recombinant expression, different downstream processing of the disulfide-rich anti-tumor peptide Ranpirnase and its effect on the growth of human glioma cell line SHG-44

Ranpirnase (Onconase) is a frogspawn-derived disulfide-rich peptide with ribonuclease activity that may be used for tumor treatment. In the present study, we established an efficient approach for preparing mature ranpirnase which may be used for research and therapeutic purposes. The designed ranpirnase precursors carried a 6xHis-tag and were recombinantly expressed in Escherichia coli. After S-sulfonation, the precursors were purified by immobilized metal-ion affinity chromatography. Following removal of the tag by aminopeptidase cleavage, cyclization and in vitro oxidative refolding, the mature ranpirnase was obtained with considerable yield, and the yield of mature ranpirnase was similar to 50-60 mg per liter cultures. In addition, ranpirnase inhibited the growth of human glioma cells SHG-44 in a dose-dependent manner. Thus the present study has provided an efficient approach for the preparation of active ranpirnase and its analogues for future studies.